| Phellinus igniarius is a precious medicinal mushroom.The fruiting body of P.igniarius,which has various medical efficacy,including enhancing immunity,anti-lipid peroxidation,anti-angiogenesis,anti-cancer,contains many kinds of compounds such as polysaceharides,flavonoids,triterpenoids,larch mushroom acids and so on.It is difficult to form fruiting body of P.igniarius in nature because of its high requirement of growing environment.In addition,the amounts of wild Phellinus igniarius were decreased sharply due to the artificial acquisition recently.Though artificial cultivation can be used to obtain fruiting body,it can only meet the needs of the part of the medicinal fungus.While the artificial cultivation technology for P.igniarius is not mature yet.Gaining mycelium by liquid fermentation,which is not affected by the season and possesses a short cycle,has broad application prospect.In order to get lots of Phellinus mycelium and actived P.igniarius flavonoids,the paper has established a liquid synthesis system for P.igniarius flavonoids on the base of the fermentation kinetics by use of response surface design.This thesis is divided into five parts,which respectively are the cognition of Phelligrins;the affect to Phelligrins production of abiotic inducer and the analysis of precursor substance by fermentation kinetics;the foundation of liquid synthesis system for Phelligrins;the extracting flavonoids with two-phase fermentation system and the exploration for functions of Phelligrins flavonoids.The main results are as follows:1.The study of chemical property of P.igniarius flavonoidsThe experiment analyzed the ethanol extracts of Phellinus fermentation broth strictly and the identification by combining with thin layer chromatography.We finaly proved that the ethanol extracts of Phellinus fermentation broth were flavonoids in according to the relevant color reaction.2.The study for fermentation kinetics of the production and activity of flavonoids after the adding precursor substance and metabolic stimulatorBy exploring the effect on the production of flavonoids and antioxidant activity(TEAC,Trolox equivalent antioxidant capacity)by adding naphthylacetate(NAA)and coumarin in the medium,the adding of NAA and coumarin was found to have a protive effect on the synthesis of flavonoids and TEAC.Moreover,it described the dynamic variation of cell growth,substrate consumption and product formation after the supplementary of NAA and coumarin in a batch fermentation system.All parameters in the model were derived from the minimum prediction model through a series of batch fermentation.The experimental data obtained after the supplementary of NAA and coumarin compared with the predicted value by the model and did the estimate of variance between them.The result presented that fermentation data inosculated with the predicted value well.3.The foundation of liquid synthesis system for Phelligrins flavonoidsThe liquid synthesis system for Phelligrins flavonoids was established by response surface design on the basis of the pre-lab experiments.At the same time,the optimal medium composition had been obtained,too.On this condition,the production of Phelligrins flavonoids reached 5.9 g/L.The process of the build of the model was as follows:first,selected the carbon,nitrogen and inorganic species as well as the optimal dosage in favor of production and activity of Phelligrins flavonoids by the single factor experiment;then screened in the 8 chemical factors which might affect Phelligrins fermentation and the production of Phelligrins flavonoids by use of PB experiment,and finaly determined coumarin,phenylalanine and Tween 80 as the most significant factors;afterwards,determining the optimal concentration of each screening factor by the central test of response surface design when the total flavonoid content and antioxidant activity of flavonoids had reached the maximum value.The significance of independent variables and the interactions between factors in the model were both determined by the estimate of variance and T or F-test.The yield of active flavonoids by use of liquid fermentation was far more than that of getting from plant tissue,which provided a theoretical basis for large-scale production of the active Phelligrins flavonoids.4.The function verification of Phelligrins flavonoidsThis study used D-galactose to form mouse senium model,and gavaged administration with Phelligrins flavonoids 30 days.At last,we detected the mouse serum super oxide dismutase(SOD),glutathione peroxidase(GSH-Px)activity and malondialdehyde(MDA),and other biochemical parameters,to explore the anti-aging of Phelligrins flavonoids.The results showed that:Compared with the model group,the test groups with different doses of Phelligrins flavonoids all had anti-aging at different levels,of which high and middle dose group had significantly enhanced in vivo peroxidase activity.The construction of diabetic mouse model took advantage of the disposable intraperitoneal injection of 100 mg/kg of streptozotocin.We monitored the fasting blood glucose levels to explore the impact of Phelligrins flavonoids to blood glucose levels in diabetic mice.The results showed that:the different doses of Phelligrins flavonoids all made lower blood glucose levels than the model group.The blood glucose levels of the mices in the high and middle dose group were back to the normal size,and the blood glucose levels of the mices in the low dose group decreased,but higher than the normal size.It indicated that Phelligrins flavonoids could reduce the blood glucose and its virtue depended on the doses.5.Preliminary extraction and purification of Phelligrins flavonoidsIn this paper,the flavonoids adsorpted by the polymer in the fermentation broth were regarded as the detached object,which was eluted by different levels of methanol,and separated into two parts ultimatly. |
